Rett syndrome is caused by mutation in the gene MECP2, encoding an epigenetic factor that binds to methylated DNA throughout the mammalian genome. MeCP2 had multiple described functional domains, but the majority of the molecule is an inherently disordered protein, meaning that is does not encode a defined secondary structure. At least two alternatively spliced isoforms and multiple phosphorylation sites of MeCP2 have been described. In addition, multiple interacting proteins of MeCP2 have been observed that are beginning to explain how MeCP2 may have such a diverse array of functions, including global chromatin structures, transcriptional repression, and transcriptional activation. Multiple gene targets of MeCP2 have emerged and help to explain how MeCP2 modulates neuronal activity and maturation, but MeCP2 is also an abundant nuclear protein that binds and acts globally on chromatin dynamics in neurons. The molecular complexities of MeCP2 appear to be consistent with a role as a master epigenetic regulator that both regulates and is regulated by multiple signal transduction and epigenetic pathways in the developing and mature mammalian brain. Therefore, understanding the post-translational modifications, isoforms and different interacting partners of MeCP2 as well as their binding sites genome-wide are expected to be critical for understanding its functions and their relevance to RTT and related neurodevelopmental disorders. This application focuses on the most abundant yet understudied MeCP2 isoform, MeCP2e1. Preliminary results demonstrate that mice selectively deficient in MeCP2e1 exhibit several features common to other Rett mouse models, including a delayed postnatal onset of neurological symptoms, reduced sociability, deficiencies in the elevated plus maze, and early lethality. The overall objective is to understand the structural and biochemical bases of MeC2e1 versus MeCP2e2 functions in pre- and postnatal brain development and their relevance to behavior. Aim 1 will investigate the structural and functional differences between MeCP2e1 and MeCP2e2 in vitro and in vivo. Aim 2 will investigate developmental roles for MeCP2e1 in recognizing the dynamic DNA methylome of developing neurons through combined genomic approaches. Aim 3 will investigate the social, behavioral, and metabolic effects of MeCP2e1 deficiency. The results of these studies are expected to be significant for understanding neurodevelopmental epigenetic pathways relevant for Rett syndrome and other more common neurodevelopmental disorders, including autism. PUBLIC HEALTH RELEVANCE: Rett syndrome is a debilitating neurodevelopmental disorder caused by mutation in the X-linked gene MECP2. While MECP2 mutations primarily cause RTT, MeCP2 is also emerging as a key modulator of disease in a number of other neurodevelopmental disorders. Therefore, understanding the basic structural and functional changes that MeCP2 undergoes in its role as a master epigenetic regulator in the developing brain will be critical for designing future therapies that may target not only RTT, but also relate neurodevelopmental disorders with higher prevalencies, such as autism, schizophrenia, and epilepsy. In this way, RTT is considered a Rosetta's stone for decoding the complexities of epigenetic pathways in human neurodevelopment in health and disease.